Mechanically competent scaffold for ligament and tendon regeneration

ABSTRACT

A multi-region device for repair, regeneration or reconstruction in articular tissue injury such as torn ligaments and tendons is provided. The device comprises at least one degradable material and biocompatible non-degradable polymeric fiber-based material, in a three-dimensional braided scaffold. The two end sections are designed for attachment of the device at the site of implantation and are designed to allow bone cell ingrowth, and one or more middle regions are designed to allow ligament or tendon cell ingrowth.

CROSS REFERENCE TO RELATED APPLICATIONS

This application is a continuation of U.S. Ser. No. 12/649,913, filedDec. 30, 2009, entitled “Mechanically Competent Scaffold for Ligamentand Tendon Regeneration”, by Cato T. Laurencin, Mark T. Aronson andLakshmi Sreedharan Nari, which claims priority under 35 U.S.C. 119 toU.S. Ser. No. 61/180,732 filed May 22, 2009, and U.S. Ser. No.61/181,033 filed May 26, 2009, all of which are herein incorporated intheir entirety by reference.

FIELD OF THE INVENTION

The present invention is in the field of implantable medical devices andprosthesis, particularly, devices useful as both a structural prostheticfor articular tissue and an in viva scaffold for the regeneration ofarticular tissue, including ligaments and tendons, and methods of makingand using the devices.

BACKGROUND OF THE INVENTION

Reconstructive surgery is based upon the principle of replacingdefective tissues with viable, functioning alternatives. In orthopaedicreconstruction, surgeons often replace damaged tissue resulting fromtrauma, pathological degeneration, or congenital deformity withautogenous grafts. The grafting of bone in skeletal reconstruction hasbecome a common task of the orthopaedic surgeon with over 863,200grafting procedures performed each year in the U.S. There are over1,000,000 procedures of various types for cartilage repair performedeach year and there are approximately 200,000 to 250,000 procedures forligament repair performed per year (Langer and Vacanti, Science, 260(5110):920-6 (1993)). Currently, autografts (tissue taken from thepatient) and allografts (tissue taken from a cadaver) are the mostcommon replacement sources for the treatment of musculoskeletal problems(Friedman, et al. Clin. Ortho., 196:9-14 (1985); Jackson, et al. Amer.J. Sports Med., 18 (1):1-10 (1990); Gazdag, et al. J. Amer. Acad Ortho.Surg., 3 (1):1-8 (1995); Shino et, al. J. Bone and Joint Surg., 70(4)1:556 (1988) and Jackson, et al. Arthroscopy, 10:442-52 (1994)). Inrepair of ligament injuries, such as injury of the anterior cruciateligament (ACL), a segment of the patellar tendon is frequently used(Jackson, et al. Amer. J. Sports Med., 18 (1):1-10 (1990)).Transplantation of autogenous grafts has been the current treatment ofchoice for cartilage and bone repair.

However, there are various problems associated with these treatments.For example, for autogenous tissue, key limitations are donor sitemorbidity where the remaining tissue at the harvest site is damaged byremoval of the graft, and the limited amount of tissue available forharvesting. Allografts are an attempt to alleviate these problems.However, this type of graft is often rejected by the host body due to animmune response to the tissue. Allografts are also capable oftransmitting disease. Although a thorough screening process eliminatesmost of the disease carrying tissue, this method is not 100% effective.Surgeons have looked to synthetic alternatives as a result of thelimitations with conventional reconstructive graft materials.

Synthetic ligament grafts or graft supports include carbon fibers,Leeds-Keio® ligament (polyethylene terephthalate), the Gore Tex®prosthesis (polytetrafluoroethylene), the Stryker-Dacron® ligamentprosthesis made of Dacron® tapes wrapped in a Dacron® sleeve and theGore-Tex® ligament augmentation device (LAD) made from polypropylene.These grafts have exhibited good short term results but have encounteredclinical difficulties in long term studies. Limitations of thesesynthetic ligament grafts include stretching of the replacementmaterial, weakened mechanical strength compared to the originalstructure and fragmentation of the replacement material due to wear.

Natural ligaments are elongated bundles of collagenous soft tissue thatserve, among other things, to hold the component bones of jointstogether. The desired characteristics for a ligament prosthesis includeappropriate size and shape, biological compatibility, capability ofbeing readily attached by the surgeon to the body of the patient, highfatigue resistance and mechanical behavior approximating that of theligamentous tissue sought to be repaired or replaced.

Ligament constructs comprising collagen fibers, biodegradable polymersand composites thereof have been developed. Collagen scaffolds for ACLreconstruction seeded with fibroblasts from ACL and skin are describedfor example in Bellincampi, et al. J. Orthop. Res. 16:414-420 (1998) andPCT WO 95/2550. A bioengineered ligament model, which includes additionof ACL fibroblasts to the structure, the absence of cross-linking agentsand the use of bone plugs to anchor the bioengineered tissue, has alsobeen described (Goulet et al. Tendons and Ligaments. In R. P. Lanza, R.Langer, and W. L. Chick (eds), Principles of Tissue Engineering, pp.639-645, R. G. Landes Company and Academic Press, Inc. 1997). U.S.Patent Application No. 20020123805 by Murray, et al. describes the useof a three-dimensional (three dimensional) scaffold composition whichincludes an inductive core made of collagen or other material, forrepairing a ruptured anterior cruciate ligament (ACL) and a method forattaching the composition to the ruptured anterior cruciate ligament(See also U.S. Patent Application No. 20040059416). WO 2007/087353discloses three-dimensional scaffolds for repairing torn or rupturedligaments. The scaffold may be made of protein, and may be pretreatedwith a repair material such as a hydrogel or collagen. U.S. PatentApplication No. 20080031923 by Murray, et al. describes preparation of acollagen gel and a collagen-MATRIGEL™ gel which is applied to a tornligament for repair of the ligament. These collagen matrices are mostlymonocomponent devices.

A number of multicomponent ligament prosthesis have been described (see,e.g. U.S. Pat. Nos. 3,797,047; 4,187,558; 4,483,023, 4,610,688 and4,792,336). U.S. Pat. No. 4,792,336 to Hlavacek, et al. discloses adevice with an absorbable component comprising a glycolic or lactic acidester linkage, and the remainder of the device comprising anon-absorbable component. The device includes a plurality of fiberscomprising the absorbable component which can be used as a flat braid inthe repair of a ligament or tendon. The required tensile strength isobtained by increasing the final braid denier. U.S. Pat. No. 5,061,283to Silvestrini discloses a bicomponent device comprising polyethyleneterepthalate and a polyester/polyether block copolymer for use inligament repair. U.S. Pat. No. 5,263,984 to Li, et al, describesprosthetic ligament which is a composite of two densities ofbioresorbable filaments. There is still a need for a device for repairof articular tissue such as the ligaments and tendons, with improvedstrength retention, load bearing capacity and ingrowth of new tissue.

It is an object of the present invention to provide a biocompatibledevice for repair, regeneration or reconstruction in articular injurywhich provides both mechanical and structure repair as well as forms ascaffold for ingrowth of cells to foam new tissue.

It is still another object of the present invention to provide a methodfor producing a device for repair, regeneration or reconstruction ofarticular injury which results in improved strength retention andingrowth of new tissue.

It is also an object of the present invention to provide a method forrepair, regeneration or reconstruction in articular injury whichcomprises implanting at the damaged area, a biocompatible polymericdevice which supports ingrowth of cells and formation of new tissue,while simultaneously providing mechanical and structural support.

SUMMARY OF THE INVENTION

A device comprising at least three phases for repair or reconstructionof articular tissue injury such as torn ligaments and tendons has beendeveloped. The device includes polymeric fiber-based degradable materialand biocompatible non-degradable polymeric fiber-based material, in athree-dimensional braided scaffold. In a preferred embodiment the deviceis composed of three regions; two end sections designed for attachmentof the device at the site of implantation, which allow for bone cellingrowth, and a middle region which serves as a scaffold for ligament ortendon cell ingrowth, resulting in the replacement of a ligament ortendon. In this embodiment, the middle region differs from the twoend-regions in size, braiding angle, porosity and/or polymercomposition. The degradable material is designed to degrade after aperiod of about nine to twelve months, to allow for repair oraugmentation of the ligament prior to the device losing the structuraland mechanical support provided by the degradable material. The deviceis made using a process of braiding degradable and non-degradablepolymeric fibers using a three-dimensional rotary braiding method or rowand column method. In the preferred embodiment, the degradable materialis poly(L-lactic acid) (PLLA) fiber and the non-degradable material is apolyester fiber.

Damaged articular tissue is repaired, regenerated, reconstructed, and/oraugmented by implanting the device at a site of injury either duringopen surgery or arthroscopically. The two ends are secured into drilledbone tunnels using interference screws, rivets, or other attachmentdevices such as sutures. Tom or damaged ligament or tendon, or allografttissue, may be sutured to or placed adjacent to the device to enhancehealing or augmentation. The scaffold can also be modified for repair ofother connective or articular injury, or other tissues.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a perspective view of the multi-region ligament repair device.

FIG. 2A is a perspective view of the bone attachment end of the devicein FIG. 1. FIG. 2B is a perspective view of the ligament tissue scaffoldcenter region of the device.

FIG. 3 is a perspective view of the end of the bone attachment end ofthe device shown in FIG. 2A.

FIGS. 4A and 4B are prospective views of the implantation of the deviceto replace a torn ACL.

DETAILED DESCRIPTION OF THE INVENTION

In the preferred embodiment, the device is used in the repair,regeneration, reconstruction or augmentation of articular injuries.Articular injuries include both intra-articular and extra-articularinjuries. Intra-articular injuries involve, for instance, injuries tomeniscus, ligament and cartilage. Extra-articular injuries include, butare not limited to, injuries to the ligament, tendon or muscle. Theseinclude injuries to the Anterior cruciate ligament (ACL), Lateralcollateral ligament (LCL), Posterior cruciate ligament (PCL), Medialcollateral ligament (MCL), Volar radiocarpal ligament, Dorsalradiocarpal ligament, Ulnar collateral ligament, Radial collateralligament, meniscus, labrum, for example glenoid labrum and acetabularlabrum, cartilage, and other tissues. The injury being treated may be,for instance, a torn or ruptured ligament. A ligament is a short band oftough fibrous connective tissue composed of collagen fibers. Ligamentsconnect bones to other bones to form a joint. A torn ligament is onewhere the ligament remains connected but has been damaged causing a tearin the ligament. The tear may be of any length or shape. A rupturedligament is one where the ligament has been completely severed providingtwo separate ends of the ligament. A ruptured ligament may provide twoligament ends of similar or different lengths. The rupture may be suchthat a ligament stump is formed at one end. The repair of the damagedtissue is achieved using the partially degradable polymeric device incombination with mechanical devices, such as interference screws,sutures and anchors

I. Ligament or Tendon Repair Device

A multi-component polymeric device serves as a template for tissueregeneration as well as provides initial structural and mechanicalsupport. The preferred ligament or tendon repair device 10 is shown inFIG. 1, based on hierarchical design methodology. The device is made upof degradable polymeric fiber and one or more non-degradable or veryslowly degradable polymeric fiber materials. In a preferred embodimentthe device is composed of three regions, with two end sections 14 a, 14b designated for attachment of the device to the bone (shown in FIGS.4A, 4B), and a middle region 12 which serves as the replacementarticular tissue. In this embodiment, the middle region 12 differs fromthe two end regions 14 a, 14 b in size, braiding angle, porosity, and/orpolymer composition (FIGS. 2A, 2B). The initial mechanical strength ofthe middle and end regions is preferably the same except for smalldifferences caused by the extra braiding revolutions in the bonyattachment region which slightly reduces strength. Ligament cellingrowth occurs in the middle region and bone cell ingrowth occurs inthe two end regions. Over time, the device begins to degrade. The endregions preferably degrade slower than the middle region due to thehigher porosity of the middle region. The middle region may degrade atone rate, or have areas of different rates of degradation.

A. Polymeric Materials

As used herein, the degradable area is that which is replaced by cellsand extracellular matrix as the polymer degrades. As used herein, thenon-degradable component retains structural and mechanical properties asthe degradable component degrades. The degradable region preferablydegrades over a period of 9 to 12 months. The non-degradable region doesnot degrade prior to one to two years following implantation.

In some embodiments, the non-degradable material is made of abiocompatible non-degradable polymer. In other embodiments, thenon-degradable component is a slowly degradable polymer. In still otherembodiments, the non-degradable material used is a mixture of one ormore non-degradable polymers and slowly degradable polymers. In theseembodiments, the slowly degradable polymer is selected such that itsrate of degradation is slower than the degradation rate for the polymerselected as the degradable component. In a preferred embodiment,degradation of the slowly degradable polymer occurs at 1-2 years.Complete degradation may not occur.

Suitable non-degradable polymers include polyethylene, polystyrene,silicone, polyfluoroethylene, polyacrylic acid, a polyaniide (e.g.,nylon), polycarbonate, polysulfone, polyurethane, polybutadiene,polybutylene, polyethersulfone, polyetherimide, polyphenylene oxide,polymethylpentene, polyvinylchloride, polyvinylidene chloride,polyphthalamide, polyphenylene sulfide, polyetheretherketone (“PEEK”),polyimide, polymethylmethacylate and/or polypropylene. In some cases,the polymer may include a ceramic such as tricalcium phosphate,hydroxyapatite, fluorapatite, aluminum oxide, or zirconium oxide.

Suitable degradable polymers include polyhydroxy acids such aspolylactic and polyglycolic acids and copolymers thereof,polyanhydrides, polyorthoesters, polyphosphazenes, polycaprolactones,biodegradable polyurethanes, polyanhydride-co-imides, polypropylenefumarates, polydiaxonane polycaprolactone, and polyhydroxyalkanoatessuch as poly4-hydroxy butyrate, and/or combinations of these. Naturalbiodegradable polymers such as proteins and polysaccharides, forexample, extracellular matrix components, collagen, fibrin,polysaccharide, a cellulose, silk, or chitosan, may also be used.

Preferred biodegradable polymers are lactic acid polymers such aspoly(L-lactic acid (PLLA), poly(DL-lactic acid (PLA), andpoly(DL-lactic-co-glycolic acid) (PLGA). The co-monomer(lactide-glycolide) ratios of the poly(DL-lactic-co-glycolic acid) arepreferably between 100:0 and 50:50. Most preferably, the co-monomerratios are between 85:15 (PLGA 85:15) and 50:50 (PLGA 50:50). Blends ofPLLA with PLGA, preferably PLGA 85:15 and PLGA 50:50 can also be used.The preferred polymer for the non-degradable region is a polyester andthe preferred polymer for the degradable region is poly(L-lactic acid(PLLA).

The proportion of the degradable to non-degradable component is selectedsuch that the non-degradable component provides retained strengthfollowing degradation of the degradable component. In a preferredembodiment, the non-degradable component is less than 30% of the device.In a more preferred embodiment, the non-degradable component forms10-30% of the device.

B. Cell Seeding

The devices can optionally be seeded with cells, preferably mammaliancells, more preferably human cells. Alternatively, they are implantedand cells may attach to and proliferate on and within the devices.Various cell types can be used for seeding. In a preferred embodiment,for ligament and tendon replacement, the cells are either mesenchymal inorigin or capable of generating mesenchymal cells. Accordingly,preferred cell types are those of the connective tissue, as well asmultipotent or pluripotent adult or embryonic stem cells, preferablypluripotent stem cells. For repair, regeneration or reconstruction ofthe ACL ligament, it may be preferable to seed the scaffold with ACLhost cells, leaving stubs of the native ACL and implanting the devicenext to or into the stubs. However, the scaffolds can be seeded with anycell type which exhibits attachment and ingrowth and is suitable for theintended purpose of the braided scaffold. Some exemplary cell typeswhich can be seeded into these scaffolds when used for repair,regeneration or augmentation of connective tissue or other tissue typessuch as parenchymal tissues, include, but are not limited to, osteoblastand osteoblast-like cells, endocrine cells, fibroblasts, endothelialcells, genitourinary cells, lymphatic vessel cells, pancreatic isletcells, hepatocytes, muscle cells, intestinal cells, kidney cells, bloodvessel cells, thyroid cells, parathyroid cells, cells of theadrenal-hypothalamic pituitary axis, bile duct cells, ovarian ortesticular cells, salivary secretory cells, renal cells, chondrocytes,epithelial cells, nerve cells and progenitor cells such as myoblast orstem cells, particularly pluripotent stem cells.

Cells used are first harvested, grown and passaged in tissue cultures.The cultured cells are then seeded onto the three dimensional braidedscaffold to produce a graft material composed of living cells andpartially degradable matrix. Each region of the scaffold can be seeded.Osteoblasts or mesenchymal stem cells in the end region help regeneratebone, ligament. Mesenchymal stem cells in the middle region regeneratethe ligament. This graft material can then be surgically implanted intoa patient at the site of ligament or tendon injury to promote healingand repair of the damaged ligament or tendon.

Growth factors and other bioactive agents may be added to the graftmaterial. In a preferred embodiment, these include fibroblast growthfactor (FGF), vascular endothelial growth factor (VEGF), epidermalgrowth factor (EGF), and bone morphogenic proteins (BMPs). Adhesivematerials such as fibronectin and vimentin can also be added. These arepreferably added in amount ranging from 0.1 nanogram to 1 micrograms.Cell isolates (for example, from marrow cells) or biological factorsisolated from blood can also be added to the graft or placed with thegraft.

II. Methods of Manufacture

The device is prepared using standard equipment and techniques, modifiedto create multiple regions (two ends for attachment and at least one,two, three or more regions). The device is braided such that the middleregion has adequate interfibrillar space and minimized thickness topromote the in-growth of tissue.

In the preferred embodiment, PLLA and polyester fibers are braided toform the three-dimensional braided devices. The device is braided, wovenor knitted so that the structure has the desired strength and stiffnessthroughout the length of the device. In a preferred embodiment, athree-dimensional rotary braiding method or row and column method isused. Three-dimensional rotary braiding is similar to traditionaltwo-dimensional-braiding technique except that it allows for greaterflexibility of fiber placement in the braided structure by allowing forthe movement of fiber bobbins independently and selectively in a flatarray of horngears. This enables the production of fiber structuresspecifically designed for an intended end use. However, as will beunderstood by those of skill in the art upon reading this disclosure,other techniques for preparing three-dimensional braided scaffolds,including row and column methods, as disclosed can also be used.

The geometric parameters which determine the shape and fiberarchitecture of three-dimensional braids includes braiding angledistribution, yarn volume fraction, number of carriers, and braidingyarn width. The braiding pattern in turn depends on braidingmachinery/technique used. A front view of the attachment end is shown inFIG. 3. The side view is shown in FIG. 2A. This contrasts with thelooser braid of the middle region shown in FIG. 2B.

The device peak load strength range is from 500 to 3200 N, with aninitial stiffness range of 200 to 700 N/mm. The number of fibers perbundle is in the range of 10 to 60 and the number of bundles per braidis in the range of 36, but it could be as low as 16 to as great as 64.

The key to the technology is the inclusion of at least three differentregions, each with the specific pore size of the matrix to promotespecific cell ingrowth. The typical range of porosity is between 50 and70%, and pore size between 177 μm and 250 μm. In the two ends forattachment to bone, the pore size is designed to allow bone cellin-growth and the middle region is specifically designed to allowligament cell in-growth.

The device is in contrast with prior art devices such as that in U.S.Pat. No. 4,792,336 which do not have regions for attachment at each endto provide both strength and bone in-growth, and a region in the middlewhich allows for ligament or tendon cell growth and retention ofstructural function until this can be replaced by the new tissue.

The devices are typically provided in a sterile kit, such as a foil orTYVEX® package. In a preferred embodiment, the device will includesutures or whip stitching in the graft, to facilitate placement. The kitwill typically include screws, sutures, or other means for attachment.

III. Methods of Use

The device is used for repair, regeneration or reconstruction ofarticular injury, by implanting the multi-region polymeric device at asite in need of articular repair or reconstruction.

Implantation is performed using standard arthroscopic technique with twoor three portals. A guide is drilled through the tibia with overdrillusing a reamer on the tibia to tibial footprint area of the ACL,followed by insertion of the guide pin to the isometric point positionover the femur near insertion of femoral insertion of the ACL. This isfollowed by an overdrill using a standard reamer on the femoral side. ABeath pin is then often used to complete drilling and the graft is thenpassed through the tibia and the femur. The graft is then secured usinginterference screws, or through the ENDOBUTTON® type technique.Visualization can be assisted using an arthrotomy. Preferably the graftis placed without an arthrotomy for a completely arthroscopically basedtechnique.

The ends 14 a, 14 b are placed in bone tunnels and secured withinterference screws, rivets, sutures 16 or other techniques, as shown inFIGS. 4A and 4B. The device is intended to be implanted using the sametechniques currently used by surgeons to implant patella tendon grafts.

In use, an appropriate number of the devices are implanted to match thebiomechanical properties of the tissue being repaired. This permits anearly return to normal function post-operatively. The implanted devicebears applied loads and tissue in-growth commences. The mechanicalproperties of the biodegradable material of the implant slowly decayfollowing implantation, to permit a gradual transfer of load to theingrown fibrous tissue. In a preferred embodiment, the degradation ofthe biodegradable material occurs after about 9-12 months. Additionalin-growth continues into the space provided by the biodegradablematerial of the implant as it is absorbed. This process continues untilthe biodegradable material is completely absorbed and only the newlyformed tissue remains. The biocompatible non-absorbable material whichis left following degradation of the degradable material provideslong-term support for the newly formed tissue.

The following non-limiting examples are provided to further illustratethe present invention.

EXAMPLES Example 1 Preparation of L-C Ligament Braid for Sheep Study

Materials and Methods

A one-meter length of L-C Ligament braid was prepared using a manualbraiding machine to create a single fiber bundle until a total of 36fiber bundles (each containing 24 fibers) have been created. Tie off thefiber bundle to the tie-off point located at the bottom of the braidingmachine. Reduce the tension in the 36 fiber bundles. Fabricate 5-cm longsection of “tight” braid. Tie-off end of the 5 cm long section of“tight” braid using a small piece of PLLA fiber that is two fibersmeasuring about 18 inches, doubled over to be 4 fibers 9 inches long.Fabricate 3-cm long section of “open” braid with one complete braidingrevolution over the 3-cm long section. Tie-off end of the 3 cm longsection of “open” braid using a small piece of PLLA fiber that is twofibers measuring about 18 inches, doubled over to be 4 fibers 9 incheslong. Tighten. Cut off excess fiber leaving strands 1 inch long. Repeatuntil the braiding of the fiber bundles is complete at the top of thebraiding machine. This yields 1 meter of braid containing 13 to 14individual 8 cm Tiger Ligaments.

Example 2 Large Animal Study

A pilot large animal study involving 16 sheep was conducted at theUniversity of New South Wales near Sydney Australia. The study wasdesigned to evaluate, in a large animal model, the in vivo response ofthe L-C Ligament for use in tendon-bone reconstruction of an anteriorcruciate ligament. The study evaluated the in vivo performance in termsof gross reaction in the knee joint, immunological reaction,radiographic evaluation in the bony tunnels, analysis of synovial fluidfor polymeric debris, mechanical performance of the synthetic ligament,histological and pathological evaluation of the synthetic ligament inthe bony tunnels and in the intra-articular space of the knee comparedto a standard doubled over autograft tendon control at 12 weeksfollowing implantation.

Materials and Methods

Animals:

Sixteen sheep were divided into four groups: four autografts (controls),four L-C ligaments, four tigers, and four autografts augmented with aL-C ligament.

Implants:

The autograft group consists of a doubled over extensor tendon.

L-C ligament is 100% PLLA, braided, measuring 8 cm in length, 2.5×2.5cm, 4 mm cross-section.

Tiger is 83% PLLA and 17% polyester, braided, measuring 8 cm in length,2.5×3×2.5 cm, 4 mm cross-section.

Method of Implantation:

ACL reconstruction was performed in the right hind limb of the sheep. 7mm tunnels were drilled in the femurs and tibias. Titanium RCI screws(7×25 mm with an 8 mm head, Smith & Nephew) were used for fixation ofthe femoral and tibial tunnels.

Animals were inducted with an IM injection of Zoletil® for sedation.Anesthesia was achieved using Isoflurane and oxygen inhalation. Allanimals received 1 g of Cephalothin intravenously and 5 mls ofBenacillin intramuscularly after induction as antibiotic prophylaxis.They also received buprenorphine (Temgesic, 0.006 mg/Kg, SC) and 4 mlsof Carprofin (Rimadyl, SC) for pain relief prior to commencement of thesurgical procedure. Twenty milliliters of blood were taken for routinepre-operative blood work.

Autograft Harvest

A 0.5 cm incision was made on the dorso-lateral aspect over the distalpart of the metacarpals. Once the lateral extensor was completely freeof the other extensors a modified Krachow stitch was inserted into thedistal free end with #2 suture. The tendon was divided proximally usinga tendon stripper, which cuts the tendon at the wrist where it exits afibro-osseous tunnel laterally. The tendon grafts were removed andwrapped in saline soaked gauze. The harvest incision sites were closedwith a resorbable 3-0 suture (Davis & Geck, North Ryde, Australia). Thegrafts will be kept moist with saline at all times during preparation. AKrackow stitch was placed at the other end of the grafts using #2 suture(Ethibond, Johnson and Johnson, North Ryde, NSW Australia).

Reconstruction

An infero-medial para-patellar incision was made. Dissection wascontinued down to the joint capsule, which was divided longitudinally.The skin edges and joint capsule were detracted to reveal theintra-articular fat pad typical of the ovine knee. The fat pad waspartially excised to reveal the cruciate ligaments. The ACL was excisedby division at its insertion into the tibia and femur, taking care notto damage the PCL. Cutting diathermy was used to excise the fat pad, butthe ACL was excised with a knife to minimize intra-articular thermalinjury. The attachment sites of the ACL to the tibia and femurwearenoted and marked at this point.

The bone tunnels were made with a 4.5 mm cannulated drill bit over aguide followed by a 7 mm canulated drill. The tibial side was drilledfrom the medial border of the tibia, just distal to the tibialtuberosity. The guide was drilled first, exiting in the joint at thecenter of the tibial insertion of the ACL. The femoral side was drilledfrom the intra-articular surface outward. The tunnel was made from theposterior part of the femoral ACL insertion with the drill guide andcannulated drill as above. The tunnel exits proximally on the lateralsurface of the lateral femoral condyle. The joint was thoroughlyirrigated with saline. The graft was placed through the tibial tunnelinto the knee joint and then into the femoral tunnel. Tension (˜40N) wasapplied to the grafts during placement of the screws adjacent to thebone. The surgical sites were closed in layers with 3-0 Vicryl (Johnsonand Johnson, North Ryde, Australia) and the skin closed with 3-0 Dexon(Davis & Geck, North Ryde, Australia). The sites were cleaned andsprayed with Op-site spray.

Post-Operative Monitoring and Recovery

Sheep were free to mobilize in their pen and fully weight bear. Nosplints were used at any time. The sheep were monitored daily in thefirst week and weekly thereafter for signs of swelling, infection,bleeding, haematoma or general ill health.

Assessments of Clinical Function

Clinical assessment of laxity and lameness at the time of surgery werequalitatively assessed based on a lameness score. A score of 0represents a lame animal that cannot load bare; 1 slight load bearing; 2partial load bearing and 3 full load bearing. Clinical assessments weremade on weeks 1, 4, and 8 weeks as well as before euthanasia at 12, 26and 52 weeks.

A functional test of anterior draw was performed at the time ofsacrifice to assess overall laxity in the same manner as the postoperative evaluation.

Animals were killed via lethal injection using Lethobarb at thedesignated time point. The right hindlimbs were harvested forexperimental endpoints. The left distal femur was harvested for testingof the contralateral limb. Twenty mls of blood was taken for routineblood work at euthanasia. This blood work was used to confirm overallanimal health status.

A post-mortem analysis was performed on the internal organs (heart,liver, kidney) as well as lymph nodes. These tissues were photographedand a portion sharply dissected and placed into phosphate bufferedformalin for routine histology. Paraffin sections were stained with H&Eand examined under normal and polarized light for histologicalappearance and the presence of any polymeric wear debris.

An anterior draw test was performed after sacrifice to examine overallknee stability with the knee in 30 degrees of flexion. The knees weregraded as lax or stable.

Mechanical Testing

Mechanical testing was performed at 12, 26 and 52 weeks as well as attime zero on cadaver knees with new devices using the reported techniqueof (Rogers et al., 1990; Walsh et at, 2007). Mechanical testing wasperformed on the femur-graft-tibia complex. The right and left hindlimbs were tested using a calibrated servo-hydraulic testing machine(MTS 858 Bionix, MTS Corporation, MN). The femur and tibia with the kneecapsule intact were placed into a drill template jig for preparation formounting. This is done to avoid any rotation of the femur or tibiaduring drilling and provide a reproducible test set-up. Two drill holesare placed in the diaphysis of the femur and tibia based on the mountingtemplate to ensure reproducible placement of the samples. The mountingtemplate oriented the samples at 45 degrees of flexion to enable theload and displacement properties to be measured in an anterior drawloading profile.

The knees were dissected following drilling of the mounting holes. Thecapsule was carefully reflected using an anteromedial incision andplaced in formalin for histological analysis described in the histologysection. The medial and lateral menisci were removed with meticulousdissection taking care not to damage the intra-articular graft. Themenisci were placed in formalin for subsequent histology. The status ofthe articular cartilage on the femur and tibia were noted andphotographed. The dissection was completed after the mounting holes weredrilled. This enabled reproducible orientation of the femur and tibiawithout any rotation or translation.

The samples were tested in anterior draw orientation (45 degrees) usinga preconditioning profile of 10 cycles followed by a stress relaxationperiod prior to testing to failure at 50 mm/minute. The peak load,energy to pull out and linear stiffness and failure mode and site weredetermined for all samples. Mechanical data was analyzed using analysisof variance followed by a Games Howell Post Hoc test with SPSS forWindows.

Synovial Fluid Analysis

Synovial fluid in the synthetic ligament group was analyzed for evidenceof polymeric debris at the time of harvest. The fluid present wascollected with an 18 gauge needle and a 20 ml syringe. The fluid withinthe syringe was photographed and the color noted. A sample of the fluidwas placed on a microscope slide and examined under polarized light forthe presence of any polymeric debris.

Computed Tomography

Computed tomography (CT) was performed on the right femur and tibia(tendon-bone reconstruction side). CT scans were performed perpendicularto the long axis of the femur and tibia using a Toshiba Scanner (Tokyo,Japan). Slice thickness was set to 0.5 mm for all scans. CT scans wasstored in DICOM format. The DICOM images was viewed using MIMICS(Materialise, Belgium). Three dimensional models was reconstructed basedand examined in the axial, sagittal and coronal planes.

Micro Computed Tomography

Micro Computed tomography (CT) was performed on the right femur andtibia (tendon-bone reconstruction side). CT scans was performedperpendicular to the long axis of the femur and tibia using a InveonScanner (Siemens, USA). Slice thickness was set to 50 microns for allscans. CT scans was stored in DICOM format. The DICOM images was viewedusing MIMICS (Materialise, Belgium). Three dimensional models wasreconstructed based and examined in the axial, sagittal and coronalplanes.

JPEG images of the DICOM images was provided in the report. Screw,tendon-bone and screw-bone interactions was qualitatively addressed in amore detailed fashion through the use of micro CT.

Magnetic Resonance Imaging

MRI was performed on the animals dedicated for histology. The right andleft hind-limbs was scanned using a 3T MRI intact prior to dissection.MR images was analyzed for signal intensity of the intra-articularportion of the graft to assess for overall status of healing compared tothe contralateral side using T2 weighted images.

Histology Right Tibia and Femur

The right tibias were processed for histology by fixation in coldphosphate buffered formalin for a minimum of 72 hours with 2 changes offormalin. Following adequate fixation, the tibias were roughly cut backwith a hack saw to isolate the tibial tunnels and the surrounding bone.The isolated bone tunnels will be dehydrated in ethanol and embedded inPMMA. Polymerized blocks were cut perpendicular to the long axis of thebone tunnel in 5 mm blocks using a Buelher saw. The blocks wereFaxitroned, polished and examined for bone ingrowth and integration withthe synthetic ligament using an environmental electron microscope(Hitachi TM1000). The screw-graft and graft-bone interface were examinedfor evidence of ingrowth and interaction as well as adverse events suchas bone resorption.

A final section from each block were cut for routine light histologyusing a Leica SP 1600 Microtome. The histological interface between thegraft, screw and tunnel will be evaluated in terms of bony response oradverse reactions in any.

The intra-articular portion of the right and left knees were harvestedand the femoral insertion marked with 7-0 suture. These samples wereprocessed in PMMA following alcohol dehydration. The blocks weresectioned using a Leica microtome. Sections were stained with H&E andTetrachrome to evaluate the new tissue infiltration. The cellularpopulation and degree of infiltration were compared versus time. Thehistology was compared to the intact left non-operated sides at eachtime point.

The medial and lateral capsular tissues from the right and left kneeswere processed for routine paraffin histology and H&E staining. Thesesections were compared to determine if the presence of the graft or thesurgical procedure resulted in a change in the synovium.

A sample of the articular cartilage from the medial femoral condyle andmedial tibial will be harvested. The cartilage samples will be fixed informalin and decalcified in formic acid. Sagittal sections will beembedded in paraffin and sections stained with H&E and Saf O to examineany changes in the articular cartilage due to surgery or the presence ofthe synthetic graft.

A sample of the medial menisci from the right and left knees wereharvested. The samples were fixed in formalin and processed for paraffinhistology. The sections were stained with H&E to examine any changes inthe menisci due to surgery or the presence of the synthetic graft.

Results

A key objective of the study was to examine the initial response of atraditional ACL autograft repair compared to an ACL reconstruction usingthe L-C Ligament™, a 100% biodegradable synthetic graft. Using an adultsheep model, the study examines numerous in vivo performance factors atvarious time horizons. The endpoints evaluated in the study includepost-operative recovery, clinical assessment of animal laxity andlameness, gross reaction in the knee joint, immunological reaction,radiographic evaluation in the bony tunnels, analysis of synovial fluid,as well as histological and pathological evaluation of the L-CLigament™, in the bony tunnels and in the intra-articular space of theknee compared to a standard doubled over (four stranded) autografttendon control at various time points following implantation.

Following initial implantations, the test animals were clinicallyassessed and monitored for lameness and load bearing ability on a fixedschedule. All subjects treated with synthetic grafts showed superiorrecoveries as compared to those treated with autografts. This is, inpart, due to the lack of harvest site morbidity as seen in the autograftmodel. Study animals experienced partial load bearing during the first 1to 3 days and had increased load bearing thereafter. When the initialsacrifices occurred, all animals had a normal gait based on aqualitative lameness score.

Approximately 12 weeks after implantation, several study animals weresacrificed for a macroscopic examination as well as histological,radiographic and pathological evaluation. A post-mortem analysis wasperformed on the key internal organs as well as the lymph nodes. Therewas no indication of any adverse events.

Radiographic imaging analysis of the reconstructed limbs, which includedx-rays and magnetic resonance imaging (MRI), confirmed the placement ofthe fixation screws after 12 weeks. A coronal view of the micro computedtomography (micro CT) at 12 weeks displayed two titanium interferencescrews and the L-C Ligament™ synthetic graft. The screws were wellplaced in the femora and tibial tunnels, and a new interface along themargins of the tunnels were noted on the CT scans, suggesting a healingbone-tunnel interface was in progress at 12 weeks following surgery. Themicro CT images also demonstrated that there was no widening of thefemoral or tibial bone tunnels. This was a positive and importantfinding.

Macro dissection of the L-C Ligament™ study animals showed no sign ofadverse reaction in the knee joint and the synovial fluid was clean andfree of debris. Furthermore, at 12 weeks, animals implanted with thesynthetic ligaments had fibrous tissue completely encompassing the graftat the intra-articular zone. Cross-sections of the synthetic grafts alsorevealed visible tissue inside the L-C Ligament™, both in theintra-articular area and on the tibial surface. These macro findings areall significant signs of healing and are indicative of a strongregenerative response.

A detailed histology of the animals sacrificed at 12 weeks confirmed themacroscopic findings. The intra-articular portion of the L-C Ligament™graft, and distal and proximal tibia were harvested, dehydrated andembedded in poly methylmethacrylate (PMMA), stained with methyleneblue/fuschin, and analyzed by light microscope. Sections were cutperpendicular to the long axis of the screw through the femoral tunnel.The intra-articular portion was sectioned perpendicular to the long axisof the graft, and the tibial tunnels were sectioned in the proximal aswell as mid portion of the tunnel perpendicular to the long axis of thescrew.

The histological study of the L-C Ligament™ animals all showed strongevidence of ligamentization in the intra intra-articular area as well asfibrous tissue integration in the entrance of the tibial bone tunnel(the fixation point). The process of ligamentization was characterizedby integration and remodeling new tissue into the intra-articularportion of the graft. Historically, tissue infiltration at the point offixation has been the major shortfall of many similar research efforts.

The synthetic graft was easily seen in sections perpendicular to thelong axis of the screw through the femoral tunnel, when viewed underpolarized light at the 12 week time point. The graft appeared intactwith no macroscopic evidence of degradation. The graft was well fixed inthe femoral tunnel and thickening of the cancellous bone at the marginsof the drill and graft was evident. New fibrous tissue was foundintegrating into the graft. Multinucleated cells were present along thesoft tissue-bone interface. Under higher magnification, plumpfibroblastic cells and new connective tissue were seen in the new softtissue integrating into the device. No evidence of resorption on thePLLA was found at 12 weeks, and the PLLA fibers appeared intact.

The intra-articular portion, sectioned perpendicular to the long axis ofthe graft, revealed the presence of the PLLA fibers with new fibroustissue integrating into the spaces present between the fibers. Highermagnification demonstrated similar fibrous tissue integration into thegraft and the presence of fibroblastic cells as noted in the bonetunnels.

Sections of the proximal portion of the tibial tunnels were alsoanalyzed at the 12 week time point. These sections were in the bonetunnel, however, the screw was not present in these sections, indicatingthe sections were above the screw. The synthetic graft was surrounded bybone when viewed under polarized light. The graft appeared intact withno macroscopic evidence of degradation and new fibrous tissue was seeninfiltrating into the graft. Under higher magnifications, the PLLAfibers appeared intact and new fibrous tissue and collagenous tissuewere found integrating into the PLLA fibers.

Sections in the graft-screw interface in the middle of the tibial tunnelwere analyzed. In these sections, the whip stitching on the graft can beseen with the screw compressing the graft against the tunnel. A closerexamination of the interface within the tibial tunnel where the screw iscompressing the graft reveals integration at the margins and stablefixation of the graft. The interface is fibrous in nature withfibroblastic cells as newly formed connective tissues.

Unless defined otherwise, all technical and scientific terms used hereinhave the same meanings as commonly understood by one of skill in the artto which the disclosed invention belongs. Publications cited herein andthe materials for which they are cited are specifically incorporated byreference.

Those skilled in the art will recognize, or be able to ascertain usingno more than routine experimentation, many equivalents to the specificembodiments of the invention described herein. Such equivalents areintended to be encompassed by the following claims.

We claim:
 1. A device for replacement or regeneration of articulartissue comprising polymeric fibers degrading at between nine and twelvemonths following implantation, and polymeric fibers not degradingfollowing implantation in less than one year, wherein bundles of thepolymeric fibers are braided into a three-dimensional braided scaffoldto form two end sections for attachment of the device at a site ofimplantation and at least one middle section between the end sections,wherein the polymer of the middle section degrades more rapidly or themiddle section is more porous than the end sections, wherein the devicehas a peak load strength range from 500 to 3200 N, with an initialstiffness range of 200 to 700 N/mm, wherein the number of fibers perbundle is in the range of 10 to 60 and the number of bundles per braidis in the range of 16 to 64, wherein the polymeric braid of the middlesection of the device has a porosity between 50 and 70%, and poresbetween 177 μm and 250 μm in diameter, and wherein the device providesarticular tissue function at the site of implantation as tissueintegrates into the device.
 2. The device of claim 1 for ligament ortendon repair, wherein the end sections are attachable to bone and allowfor ingrowth of bony tissue.
 3. The device of claim 2 wherein theporosity of the middle section allows in-growth of connective tissue. 4.The device of claim 1 produced by a three-dimensional rotary braidingmethod or row and column method.
 5. The device of claim 1, wherein themiddle section differs from the end sections in size, braiding angle, orporosity.
 6. The device of claim 1 wherein between 70 and 90% of thepolymeric fibers are degradable in less than one year.
 7. The device ofclaim 1 wherein the degradable polymer fibers are made up of polymersselected from the group consisting of poly(L-lactic acid (PLLA),poly(DL-lactic acid (PLA), poly(DL-lactic-co-glycolic acid) (PLGA),polyorthoesters, polyanhydrides, polyphosphazenes, polycaprolactones,polyhydroxyalkanoates, biodegradable polyurethanes,polyanhydride-co-imides, polypropylene fumarates, polydiaxonane,polysaccharides, collagen, silk, chitosan, and celluloses.
 8. The deviceof claim 1 wherein the non-degradable polymer fibers are made up ofpolymers selected from the group consisting of polyethyleneterephthalate (PET), polyetheretherketone (PEEK),polytetrafluoroethylene (PTFE), polyester, polyethylene, polyamide,polyimide, polyurethane, polybutadiene, polybutylene, and polypropylene.9. The device of claim 1, wherein the device is seeded with cells,ingrowth of which is supported by the scaffold.
 10. The device of claim9 wherein the cells are selected from the group consisting ofmesenchymal cells, cells generating mesenchymal cells, fibroblasts,pluripotent stem cells, and multipotent stem cells.
 11. A method forrepairing, regenerating, replacing or reconstructing a damaged tendon orligament in a patient comprising implanting at a site of a damagedtendon or ligament the device of claim
 1. 12. A kit comprising thedevice of claim 1 and means for attachment.
 13. A method of making thedevice of claim 1 comprising braiding a three-dimensional scaffoldcomprising polymeric fibers degrading at between nine and twelve monthsfollowing implantation, and polymeric fibers not degrading followingimplantation in less than one year, wherein the polymeric fibers arebraided into a three-dimensional braided scaffold to form two endsections for attachment of the device at a site of implantation and atleast one middle section between the end sections, wherein the polymerof the middle section degrades more rapidly or the middle section ismore porous than the end sections, wherein the device provides articulartissue function at the site of implantation as tissue integrates intothe device.
 14. A device for replacement or regeneration of articulartissue comprising bundles of fibers formed of polymer degradingfollowing implantation, wherein the bundles of the polymeric fibers arebraided into a three-dimensional braided scaffold to form two endsections for attachment of the device at a site of implantation and atleast one middle section between the end sections, wherein the middlesection is more porous than the end sections, wherein the device has apeak load strength range from 500 to 3200 N, with an initial stiffnessrange of 200 to 700 N/mm, and wherein the device provides articulartissue function at the site of implantation as tissue integrates intothe device.
 15. The device of claim 14 wherein the fibers in the bundlesare multifilament fibers.